The study will be concerned with protein kinase from the pig parotid and submaxillary gland and a delineation of the substrates for protein kinase in microsomal preparations as well as zymogen granules. Soluble protein kinase will be isolated from the glands according to published methods. The enzyme is being characterized kinetically in terms of substrate specificity, activation by Mg ions and ATP, stimulation by cyclic nucleotides and sensitivity to inhibition by Ca ions and protein kinase inhibitor. The isolation of this enzyme in relatively pure form will allow a more extensive investigation into the interaction of the enzyme with the substrates of interest. Phosphorylation of microsomal preparations and zymogen granules by protein kinase will be studied kinetically and a detailed characterization of the phosphorylated products will be pursued. Lipid extraction, determination of the sensitivity of the phosphorylated products to various gents, and amino acid analysis will be used in this characterization. Isolation of protein kinase substrates will be accomplished prior to amino acid analysis by polyacrylamide gel electrophoresis. Proteins kinase activity in the microsomes and zymogen granules will be tested and its kinetic characteristics compared to that of the soluble protein kinase from the same gland. Since some protein kinase-catalyzed phosphorylation events are known to increase the substrates' affinity for calcium, and others are postulated to have this effect, the effect of phosphorylation of these organelles will be determined by measuring calcium interaction with them. Calcium binding and calcium accumulation in the presence of a precipitating anion (e.g. oxalate) will be estimated as a function of the phoshorylation of the organelles. The possible content of protein kinase and phosphorylatable proteins in saliva will be evaluated as an indication of the level of secretion of these proteins from the gland during exocytosis.